A MOLECULAR BASED SEEDLING ASSAY SYSTEM TO STUDY THE GERMINATION AND PRIMARY INFECTION BY PLASMODIOPHORA BRASSICAE

M. Lewis, A. Wakeham , R. Kennedy
The life cycle of P. brassicae consists (in part) of a single motile zoospore infecting a root hair cell, followed by replication within host cells before transposition to the root cortical cells where further spore formation occurs. Due to the complex nature of the lifecycle, it has been necessary to consider alternative methods to determine the viability of this organism. If DNA levels are considered at an appropriate stage of the life cycle (during the resting spore or primary zoospore stage) then the level of DNA is directly comparable to the number of spores present as these lifecycle stages have been shown to contain haploid nuclei.
The objectives of this study were to construct a seedling assay system which uses the sensitivity of molecular methods to allow the determination of the number of germinating spores as a measure of viability, so that factors affecting viability can be investigated. Employing quantitative PCR, a seedling assay has been developed that allows the germination of resting spores to be quantified thus giving an indicator of spore viability within two weeks. The potential for monitoring levels of infection resulting from resting spore germination was also investigated in relation to infection of Brassica rapa ssp. pekinensis ‘Granaat’ (Chinese cabbage). Infection of root tissues by zoospores produced from a resting spore suspension was demonstrated by quantifying the levels of P. brassicae DNA over the period corresponding to primary infection. It was found that significantly reduced levels of P. brassicae DNA were detectable across a range of resting spore suspension concentrations after incubation with the brassica seedlings, and that P. brassicae DNA could be isolated from root tissue after the incubation period which, along with microscopy, confirmed seedling infection.
Lewis, M., Wakeham , A. and Kennedy, R. (2013). A MOLECULAR BASED SEEDLING ASSAY SYSTEM TO STUDY THE GERMINATION AND PRIMARY INFECTION BY PLASMODIOPHORA BRASSICAE. Acta Hortic. 1005, 583-589
DOI: 10.17660/ActaHortic.2013.1005.72
https://doi.org/10.17660/ActaHortic.2013.1005.72
quantitative polymerase chain reaction, seedling assay
English

Acta Horticulturae